Decoupling of heat and CO2 release during decomposition of cellulose and its building blocks in soil

Abstract

The degradation of large biopolymers, such as cellulose, in soil requires several enzymatic hydrolysis steps to produce simpler substrates for microbial uptake. The synthesis of these enzymes requires energy and takes time until they are fully expressed. However, the heat release associated with enzymatic hydrolysis and the temporal delay between this initial heat release and the final carbon mineralization to CO₂ is largely unknown. In this study, we investigated the dynamics of heat and CO₂ release during the sequential decomposition of cellulose to its building blocks, cellobiose and glucose, in soil and related these processes to activities of cellobiohydrolase and β-glucosidase driving the corresponding steps of cellulose decomposition. Moreover, we estimated catabolic heat release during the stepwise enzymatic production of oligo- and monomers in soil by employing fluorogenically labeled substrates. This amounted to the absolute value of 26.5 kJ mol C⁻¹, approximately 6.5 % of the total combustion enthalpy stored in the applied cellulose.

By three complementary approaches, we confirmed that cellobiohydrolase rather than β-glucosidase is the bottleneck step of enzymatic hydrolysis. First, a 36 h temporal decoupling between the heat and CO₂ formation peaks occurred during step-wise enzymatic hydrolysis of cellulose performed by cellobiohydrolase and β-glucosidase towards final mineralization. This decoupling was not observed in the next sequential step of cellobiose hydrolysis by β-glucosidase. Remarkably, heat and CO₂ release evolved more slowly during cellulose degradation compared to that of its building blocks, cellobiose and glucose. Second, the enzyme activity of β-glucosidase more than doubled that of cellobiohydrolase during cellulose degradation. Third, heat release after the addition of flurogenically labeled substrate to soil, which mimics the steps of cellulose degradation, was faster in the step of glucose production than that of cellobiose production. This study highlights the novel mechanistic insights facilitated by calorespiroemetric monitoring of carbon decomposition at high temporal resolution.